Recently, organic solvents, and particularly halogenated hydrocarbons, have been used in large amounts as cleaners and so forth in advanced industries. Since growing attention is being focused on contamination of groundwater and soil caused by these substances or waste water containing these substances, there is a desire to implement countermeasures against this contamination immediately.
Examples of known physical methods that have been employed in the past as countermeasures include an air stripping method in which the contaminated soil is excavated and air is blown through the soil to volatilize the halogenated hydrocarbons and adsorb it with activated charcoal and so forth, and a vacuum extraction method in which the contaminated soil is pounded into a pipe after which a vacuum is drawn to aerate the soil and remove the contaminants. These methods are considered to be able to be applied to decontamination of groundwater as well.
However, these methods have the disadvantage of requiring a large amount of energy, such as for blowing in air. In addition, the former has the disadvantage of requiring that the soil be excavated, while the latter has the disadvantage of extraction efficiency decreasing as the concentration of contaminant decreases, thus making the cleaning difficult. Moreover, from the viewpoint of preventing secondary contamination such as air pollution, these methods require that separate contaminants be detoxified in order to adsorb onto activated charcoal and so forth.
On the other hand, research has been conducted in recent years on so-called biocleaning methods in which contaminants are efficiently decomposed and detoxified by microorganisms. Since these methods utilize the decomposition mechanism of microorganisms, they do not require a large amount of energy as compared with the above-mentioned physical methods. They are also able to completely decompose and detoxify contaminants without causing secondary contamination. Moreover, the cleaning can be performed even at low concentrations of contaminants, thus enabling decontamination to be performed over a wide area at the original location and creating significant expectations of low costs.
Examples of methods used to purify contaminated soil by microorganisms include a solid phase treatment in which microorganisms are mixed into excavated soil with nutrient sources such as phosphorous and nitrogen to promote decomposition of contaminants, a slurry treatment in which microorganisms are mixed into excavated soil with water and nutrient sources to treat the soil in the liquid state and promote decomposition of contaminants, and an original location treatment in which air, nutrient sources and so forth are injected into contaminated soil without excavating to promote decomposition of contaminants by microorganisms present in the soil.
Among the above-mentioned biotreatment techniques, since soil excavation is required and the application range is limited in the case of the solid phase treatment and slurry treatment method, treatment and equipment costs are relatively high.
On the other hand, the original location treatment method involves relatively low costs and allows treatment over wide area. However, the cleaning rate is slow since the the absolute number of soil microorganisms is low. In the case of compounds that are difficult to decompose such as halogenated hydrocarbons in particular, there is a possibility that microorganisms being able to decompose contaminants in the soil may not be present in the soil, thus making cleaning impossible. In this case, acquiring microorganisms that are able to decompose halogenated hydrocarbons and inoculating them into the soil enables the cleaning rate to be improved and soil to be purified even though microorganisms being able to decompose the contaminants are not present in the soil.
A halogenated hydrocarbon contaminant, trichloroethylene (TCE), is widely used in the IC industry, in dry cleaning and so forth. It is particular important as a contaminant since it is reported to be carcinogenic. Known examples of microorganisms that decompose TCE include the methane assimilating microorganisms Methyrosinus tricosiorium OB3 (Japanese Unexamined Patent Publication No. 4-501667, Japanese Unexamined Patent Publication No. 5-212371) and Methyrosinus tricosporium TUKUBA (Japanese Unexamined Patent Publication No. 2-92274 and Japanese Unexamined Patent Publication No. 3-292970),
Pseudomonas, such as Pseudomonas putida F1 (Japanese Unexamined Patent Publication No. 64-34499), Pseudomonas putida BH (Fujita, et al., Chemical Engineering, 39, (6), p.494-498, 1994), Pseudomonas putida UC-R5 and UC-P2 (Japanese Unexamined Patent Publication No. 62-84780), Pseudomonas putida KWI-9 (Japanese Unexamined Patent Publication No. 6-70753), Pseudomonas mendocina KR-1 (Japanese Unexamined Patent Publication No. 2-503866 and 5-502593), Pseudomonas cepacia G4 (Japanese Unexamined Patent Publication No. 4-502277) and Pseudomonas cepacia KK01 (Japanese Unexamined Patent Publication No. 6-296711) and other microorganisms such as Alcaliaenes eutronus JMP134 (A. R. Harker, Appl. Environ. Microbiol., 56, (4), 1179-1181, 1990), Alcaliaenes eutropus KS01 (Japanese Unexamined Patent Publication No. 7-123976), and the ammonia bacteria Nitrosomonus europaea (D. Arciero, et al., Biochem. Biophys. Res. Commun., 159, (2), 640-643, 1989) are known.
Pseudomonas cepacia KK01 in particular is reported to decompose TCE at an initial concentration of 30 ppm to a concentration of 15 ppm in liquid culture, and TCE in soil having an initial concentration of 5 ppm to a concentration of 1 ppm (Japanese Unexamined Patent Publication No. 6-296711). In addition, Alcaliaenes eutropus KS01 is reported to have the ability to decompose TCE at a concentration of 50 ppm in a liquid culture to a concentration below the detection limit, and decompose TCE in the soil at 1 ppm below the detection limit (Japanese Unexamined Patent Publication No. 7-123976).
However, when testing the decomposing abilities of these microorganisms, decomposition is demonstrated at extremely high cell concentrations (1.times.10.sup.8 cells/ml) in all cases. When considering that this concentration is unrealistic in the actual soil environment, the decomposing abilities of these microorganisms is not always considered to be high. Thus, in the case of using microorganisms for soil cleaning, the microorganisms have sufficient decomposing ability and are able to demonstrate that ability in the special environment of the soil, such as in the presence of wild microorganisms. In addition, it is preferable that the tolerance of the microorganisms to TCE, the target of decomposition, be high, and that they also have the ability to decompose dichloroethylene (DCE), which is a partial decomposition product of TCE.
Disclosure of Invention
The object of the present invention relates to microorganisms that efficiently decompose halogenated hydrocarbons, and particularly high concentrations of TCE, DCE and so forth, as well as a cleaning process for water or soil that uses those microorganisms.
The present invention provides microorganisms having the ability to decompose halogenated hydrocarbon and belonging to the genus Burkholderia, and some of those microorganisms belonging to the species Burkholderia cepacia. Examples of these microorganisms include Burkholderia N16-1 (FERM BP-5504), Burkholderia cepacia N15-1 (FERM BP-5502) and Burkholderia cepacia N15-2 (FERM BP-5503). Moreover, the present invention provides a process for cleaning water or soil where the above-mentioned microorganisms are added to water or soil containing halogenated hydrocarbon.
The present invention is also characterized by the addition of microorganism activator in combination with the above-mentioned microorganisms. These microorganisms have the ability to decompose 50% or more of 100 ppm of trichloroethylene in 2 days, or to decompose 100% of 30 ppm of trichloroethylene in 18 hours.